The PDZ Binding Motif of Human Papillomavirus Type 16 E6 Induces PTPN13 Loss, Which Allows Anchorage-Independent Growth and Synergizes with Ras for Invasive Growth

Abstract

The human papillomavirus (HPV) oncogene E6 has been shown to perform multiple functions (p53 degradation, telomerase activation, etc.) that play a role in oncogenic transformation. Beyond known E6 functions, an undefined mechanism that allows cellular invasion requires the E6 PDZ binding motif (PDZBM). Here, we show that HPV type 16 (HPV16) E6 interacts with and induces loss of a protein tyrosine phosphatase (PTPN13) in a PDZBM-dependent manner. PTPN13 loss induced either by the presence of E6 or by a short hairpin RNA strategy allows for anchorage-independent growth (AIG) and synergy with a known oncogene, Ras v12 , resulting in invasive growth in vivo. Restoring PTPN13 expression reverses AIG in cells lacking PTPN13. A genomic analysis of colorectal carcinoma has identified an association between PTPN13 loss-of-function mutations and aberrant Ras signaling. Our findings support this correlation and provide methods for further evaluation of the mechanisms by which PTPN13 loss/Ras expression leads to invasive growth, the results of which will be important for treatment of HPV-related and non-HPV cancer.Significant associative evidence supports a role for human papillomavirus type 16 (HPV16) in the carcinogenic progression of at least 25% of head and neck squamous cell cancer (HNSCC) cases (27, 38). In particular, the incidence of HNSCC of the tonsillar region is increasing (18,20,38). The epithelial surfaces surrounding the tonsil (oropharynx) are cancerous in a very high (60%) proportion of cases that are HPV positive, and these cases present with more metastatic and advanced disease states (34, 38) than HPV-negative cases. The incidence of HPV-related HNSCC is also increased in individuals with human immunodeficiency virus/AIDS disease (10). While the data presented in the following studies pertain to HPV-related HNSCC they may also be applicable to cases of HPV-related cervical cancer, which remains the second leading cause of cancer-related death in women. Therefore, understanding the viral mechanisms that lead to cancer will help meet a need for HPV-specific targeted therapy.One of the major HPV viral oncogenes that allow progression to cancer is HPV16 E6. E6 has many potential carcinogenic effects, including telomerase activation and p53 degradation (26, 35). However, an understood mechanism that potentiates malignant progression has been associated with a PDZ (PSD-95, Discs Large, ZO-1) binding motif (PDZBM), a specific amino acid sequence that mediates protein-protein interactions with corresponding PDZ domain-containing proteins. High-risk HPV virus types 16, 18, 31, and 33 possess this binding motif at the E6 C terminus. In addition to this correlative evidence of increased malignant potential in humans, mouse studies demonstrated that PDZBM is required for HPV-related malignancy (37).The studies described below explored the role of the E6 PDZBM in binding and inducing loss of a PDZ domain containing phosphatase PTPN13. PTPN13 is a member of the nonreceptor phosphatases and specifically falls into a cl...

Human papillomaviruses, cervical cancer and cell polarity

“…Interestingly, shRNA-induced PTPN13 loss in tonsillar epithelial cells will induce anchorage-independent growth, and PTPN13 loss can also cooperate with activated ras to induce invasive cell growth. Equivalent phenotypes are induced by wildtype, but not PDZ binding-defective, HPV-16 E6 (Spanos et al, 2008b). This provides compelling evidence that the PTPN13 interaction plays a role in HPV-induced malignancy, and it will be interesting to determine how this pathway might impact upon the control of cell polarity.…”

“…Like MUPP1, PSD95 (Dlg4, another Dlg family member) (Handa et al, 2007) is thought to be a scaffold for signalling complexes, but has so far mainly been described at neuromuscular synapses. The PTPH1/ PTPN3 protein tyrosine kinase (Jing et al, 2007;To¨pffer et al, 2007) and the PTPN13 non-receptor kinase (Spanos et al, 2008b) are also described as HPV E6 targets. Tyrosine kinase phosphorylation is involved in signalling and is commonly altered in transformed cells and cancers.…”

Human Papillomavirus: Biology and Pathogenesis

“…Degradation of PDZ proteins results in cellular transformation due to loss of cell-cell contact and loss of cell polarity (Storrs and Silverstein, 2007). In addition, it has been demonstrated that the degradation of phosphatase PTPN13 by E6 results in anchorage-independent growth and a Ras-dependent invasive phenotype (Spanos et al, 2008).…”

“…HR-HPVE6 proteins also bind to hScrib, a protein involved in epithelial tight junctions, mediating the adhesion of basal cells to the ECM, and at least in some cell types it has been shown that E6 mediates hScrib degradation (Nakagawa & Huibregtse, 2000). Recently, it has been demonstrated that PTPN3, a membrane-bound tyrosine phosphatase that regulates growth factor receptors, is also a PDZ protein that binds and is disrupted by E6 (Jing et al, 2007;Spanos et al, 2008).…”

Human Papillomaviruses Oncoproteins

“…Several proteins involved in apoptosis and immune evasion (Bak, FADD,c-Myc, NFX1, procaspase 8, etc) [45,46,47,48] are also targeted by E6 HPV. Bak, a member of the Bcl-2 family, is a proapoptotic protein whose interaction with the viral oncogene leads to the inhibition of apoptosis.…”